The plant Artemisia annua (family: Asteraceae) produces a sesquiterpenoid lactone endoperoxide named artemisinin which is a promising antimalarial drug effective against Plasmodium falciparum, Plasmodium vivax at nanomolar concentration. Artemisinins are active against Schistosoma mansoni and S. japonicum in-vitro and in-vivo in experiments in animals. These schistosomes, like malarial parasites, degrade haemoglobin and produce hemozoin. These compounds are also active against Leishmania major, Toxoplasma gondii and Pnenmocystic carinii in-vitro and against P. carinii in-vivo. Artemisinins have immunosuppressive activity and also potential anticancer activity. For these activities, the doses of artemisinin required are substantially higher than the dose for antimalarial activities. According to Meshnick et at., (1996) (Microbiological Reviews 6: 301‥315) the antimalarial endoperoxides including artemisinin, dihydroartemisinin and arteethers, are not likely to be useful for other therapeutic purposes except against malarial parasites.
Although artemisinin rapidly suppresses the activity of parasites like Plasmodium vivax and P. falciparum, problems with high rate of recrudescence (>10% recrudescence infections), short half life persist. Hence, there is a need to develop new drugs against quinolone resistant pathogenic bacteria. It is a known fact that clinically used antibacterial broad spectrum compounds such as quinolones which exhibit DNA gyrase activity of Mycobacterium sp. (causing tuberculosis), Haemophilus sp. and Haemophilus influenzae are gradually becoming ineffective due to the occurrence of mutatious in gyrase genes and their natural selection under continuous use of such drug.
The compound α arteether developed as antimalarial drugs by Central Drug Research Institute (CDRI), Lucknow, India and Central Institute of Medicinal & Aromatic Plants (CIMAP), Lucknow, India, after phase II clinical trial is a stable derivative of artemisinin. Earlier we have found a novel property of α-arteether as being effective against the gyr A mutant strains of E. coli but ineffective against wild type strains (U.S. Pat. No. 6,127,405). Also we have developed a strategic and novel composition comprising α arteether and nalidixic acid or quinolone drugs which is useful as an advanced generation drug to counter the resistance development itself and having a potential to be used in treating infectious diseases and in inhibiting the resistance developed due to mutation in the gyr A gene of bacteria, particularly in those cases where drug resistant strains are known to appear very frequently (U.S. Pat. No. 6,423,741).
In an earlier invention a method was also provided for maximization of artemisinin yield of the plant Artemisia annua, said method comprising sowing seeds of Artemisia annua plant on raised bed nursery during second and third week of December and maintaining the moisture throughout; transplanting seedlings thus obtained bearing at least 5-15 leaves into the main field fertilized with fertilizer, preferably NPK @ 80,40,40 kg/ha to achieve a population density of 50,000 to 200,000 per ha followed by light irrigation in the second week of March and irrigation every fortnight thereafter; harvesting the crop four times by cutting the plant tops leaving 75-100 cm part of plant for further regeneration, the said harvests are performed in a manner that the first harvest is done in fourth week of May, second harvest in third week of July, third harvest in second week of September and fourth harvest in third week of October of each year; and at each harvesting time care is taken to care at least one green branch, and extracting artemisinin from the plant tissue immediately after each harvest. (U.S. Pat. No. 6,393,763).
Considering the high value of the chemical artemisinin for use in derivatization to different semisynthetic product of immence importance the need of the hour is to still increase the yield. Agronomic practices and scheduling of the harvest timings to obtain higher biomass yield do not take into account the genotypes as all the plants are harvested together. Since the plant Artemisia annua is highly cross pollinated like the members of family Asteraceae the chemical character like ‘artemisinin content’ seggregate like any other phenotypic characters as multigenic characters always segregate in the progeny population. Due to this all the progeny plants of the high artemisinin containing plant may not yield same amount of the chemical. Some will be high, some medium and some very low.
Considering the problem of identification of the high artemisinin containing plant genotypes at the nursery stage, to discard the low artemisinin genotypes for the purpose of planting only those genotypes which could produce high amount of artemisinin during maturity in the main field a systematic approach for identification of DNA marker was launched. In this process the marker was identified which could differentiate the high artemisinin genotypes from low artemisinin genotypes at the seedling stage itself. These selected seedlings showing the presence of the DNA marker then could be taken for further matting between them to generate plants produce highest biomass as well as higher artemisinin.